The Mg2+ content of the heart is significantly reduced during acute hypoxia/anoxia and during Ca2+ deprivation (a model for acute ischemic states), suggesting that Mg2+ loss may mediate the pathophysiological changes that accompany acute myocardial oxygen deprivation. The objective of this proposal is to test the hypothesis that, in the isolated perfused rabbit heart, the intracellular Mg2+ concentrations are important in regulating the arachidonic acid content and overall fatty acid composition of phospholipids. In the isolated rabbit heart, the synthesis of prostaglandins from exogenous arachidonic acid is significantly increased during perfusion with Ca2+ free buffer, while the incorporation of labelled arachidonate into tissue phospholipids, especially phosphatidyl choline and phosphatidyl ethanolamine, is significantly reduced. The difference on prostaglandin output in response to exogenous arachidonic acid are linearly correlated with the tissue Mg2+ content, and cannot be accounted for by differences in the Km or Vmax of cyclooxygenase, changes int he permeability of the heart to arachidonate or changes in the profile or prostaglandins synthesized. Arachidonyl CoA synthetase and lysophosphatidyl choline acyl transferase (enzymes which are important in the incorporation of arachidonic acid into lipids) require Mg2+ for their activity. Thus, changes in the activities of these enzymes might be brought about by experimental manipulations which change the Mg2+ content of the tissue. Experiments are proposed to: 1) study more extensively the effects of variation in the tissue cation contents and the conversion of exogenous arachidonic acid to prostaglandins; 2) study the effects of tissue cation manipulations on the incorporation of arachidonic acid into tissue lipids; and 3) determine whether the activities of arachidonyl CoA synthetase and/or lysophosphatidyl choline acyl transferase are changed under experimental conditions (such as Ca2+ free perfusion) which result in reduced incorporation of arachidonic acid into lipids. The oxygen tension or the ionic composition of the perfusion buffer will be varied; these manipulations are expected to result in shifts in the total tissue content of Ca2+, Mg2+, Na+, and K+. Conversion of arachidonic acid to prostaglandins and the incorporation of radiolabelled arachidonic acid into neutral lipids and phospholipids will be measured. The data will be examined for significant correlations between changes in the disposition of arachidonic acid (i.e., changes in the synthesis of prostaglandins from exogenous arachidonic acid and changes in the incorporation of exogenous arachidonic acid into lipids) and shifts in the ion content of the tissue. The activities (Km and Vmax) of arachidonyl CoA synthetase and lyosphosphatidyl choline acyl transferase will be measured in hearts perfused with and without Ca2+. The effect of added Ca2+ and/or Mg2+ on the activities of these enzymes will also be measured.